Replication of biotinylated human immunodeficiency viruses.

Area of Science:

• Virology
• Molecular Biology
• Biochemistry

Background:

• The Escherichia coli biotin ligase BirA-biotin acceptor sequence (BAS) system was previously adapted for human immunodeficiency virus type 1 (HIV-1).
• Biotinylation of HIV-1 integrase (NLXIN(B)) and matrix (NLXMA(B)) proteins was achieved.

Purpose of the Study:

• To construct a human immunodeficiency virus (HIV) permissive cell line stably expressing BirA (SupT1.BirA).
• To evaluate the replication and biotinylation efficiency of HIV-1 and HIV-2 constructs in the SupT1.BirA cell line.
• To demonstrate the utility of the biotinylation system for labeling and capturing HIV protein complexes.

Main Methods:

• Construction of a human immunodeficiency virus (HIV) permissive cell line (SupT1.BirA) stably expressing BirA.
• Generation of HIV-1 viruses with biotinylated integrase (NLXIN(B)) and matrix (NLXMA(B)) proteins.
• Construction of three HIV type 2 (HIV-2) viruses with BAS inserted into vpx and vpr accessory genes.
• Assessment of viral replication and protein biotinylation in SupT1.BirA cells.

Main Results:

• NLXMA(B) HIV-1 replicated similarly to wild-type levels, with biotinylated Gag and MA proteins observed.
• NLXIN(B) HIV-1 replication was severely reduced.
• Three HIV-2 viruses with BAS insertions in Vpx and Vpr were replication-competent.
• HIV-2 target proteins were efficiently biotinylated and incorporated into virions.

Conclusions:

• The SupT1.BirA cell line supports the replication of biotinylated HIV-1 and HIV-2 viruses.
• The BirA-BAS system is effective for in vivo biotinylation of HIV proteins, including accessory proteins.
• This biotinylation strategy holds potential for labeling and capturing HIV protein complexes within replicating viruses.